The polyamines, putrescine, spermidine, and spermine, are major polybasic compounds in all living cells. These amines are important for many systems related to growth and differentiation. For many years we have been studying how these polyamines are synthesized, how their biosynthesis and degradation are regulated, their physiologic functions, how they act in vivo, and the structure of the various biosynthetic enzymes. For this purpose we have constructed null mutants in each of the biosynthetic steps in both Escherichia coli and Saccharomyces cerevisiae, and have prepared overexpression systems for the biosynthetic enzymes. Our overall studies have aimed at the use of these mutants to elucidate the physiological functions of the polyamines.[unreadable] [unreadable] Our recent studies have been concerned with measuring the internal concentration of polyamines in Saccharomyces cerevisiae under various growth conditions, and, in particular, the internal concentration found when auxotrophic mutants are grown on limiting spermidine. These studies required modification of our analytical techniques to permit a more sensitive determination. We have found that the internal concentration of spermidine needed for growth is much less than the large amounts normally present in the cells, and that this lower concentration of spermidine is largely used for the biosynthesis of hypusine, a modification of eIF5A that is critical for protein biosynthesis. These studies are of particular significance since they demonstrated that this modification of eIF5A is the most critical function for spermidine; i.e. both for protein biosynthesis and growth. [unreadable] [unreadable] We have also been interested in investigating why the yeast cells (and other cells) normally contain the higher concentration of polyamines; i.e. much more than needed for growth. We have shown that one system requiring the larger amounts of spermidine and spermine is the biosynthesis of beta-alanine and pantothenate.